羊水来源多潜能干细胞的培养鉴定及其定向分化能力研究

Cultivation,identification and directional differentiation potency of human amniotic fluid-derived multipotential stem cells

目的建立人羊水来源多潜能干细胞的体外培养体系,探讨其生物学特性和定向分化能力。方法取人孕中期羊水标本进行原代培养,采用贴壁法分离获得干细胞。以流式细胞仪检测细胞表面抗原的表达,并使用特定培养基诱导羊水多潜能干细胞向成骨细胞、脂肪细胞、神经细胞分化。结果体外培养的羊水干细胞呈纺锤形,增长迅速,细胞表达Oct-4、SSEA-4、CD29、CD44、CD105、CD117,但不表达造血干细胞表面标志CD45、CD133。羊水干细胞在体外诱导可以分化出成骨细胞、脂肪细胞和神经细胞。结论分离获得的干细胞体外增殖能力强,可表达多潜能干细胞和胚胎干细胞的标志基因,符合多潜能干细胞的特点。在适宜的培养条件下,羊水多潜能干细胞具有向成骨细胞、脂肪细胞、神经细胞分化的能力。

Objective To establish a procedure for culturing amniotic fluid-derived multipotential stem cells in vitro,and study their biological characteristics and differentiation potency.Methods Amniotic fluid-derived stem cells were isolated from second-trimester amniotic fluid by adherence.The phenotypes of amniotic fluid-derived stem cells were detected by flow cytometry.The amniotic fluid-derived stem cells were induced to differentiate into osteoblasts,adipocytes and neurocytes by specific culture medium.Results The stem cells were successfully isolated from 8 of 10 second-trimester amniotic fluid samples.The amniotic fluid-derived stem cells were spindle in shape and proliferated rapidly.Primary cultured cells grew slowly,and then the growth became rapid following passaging.The chromosome karyotype of the 20th passage was normal,remaining to be 46,XX or 46,XY.These cells shared immunophenotype of both multipotential stem cell and embryonic stem cell markers,as they were positive for Oct-4,SSEA-4,CD29,CD44,CD105 and CD117 expressions,and negative for CD45 and CD133 expressions.Amniotic fluid-derived stem cells could differentiate into osteoblast,adipocyte and neurocyte in vitro by specific culture medium.Conclusions The stem cells isolated from second-trimester amniotic fluid have a great potential of proliferation,they may express both embryonic and multipotential stem cell markers,and they are in accordance with that of multipotential stem cells.Amniotic fluid-derived stem cells have the ability to differentiate into osteoblast,adipocyte and neurocyte in suitable condition.